Optimization studies for the bioconversion of Jerusalem artichoke tubers to ethanol and microbial biomass

Summary A total of 8 yeast and microbial cultures have been grown in the extract derived from the tubers of Jerusalem artichoke (Helianthus tuberosus) and screened according to the following optimization criteria: rates and yields of ethanol production, rates and yields of biomass production, and % of original sugars utilized during fermentation. Batch growth kinetic parameters were also determined for the cultures studied.     

Adsorption and separation of proteins by a smectitic clay mineral

The adsorption of proteins by a smectitic clay mineral was investigated. The clay used in this study is a mixture of montmorillonite and amorphous SiO₂. Due to the high porosity the montmorillonite units are accessible for protein adsorption. The amorphous silica prevents the montmorillonite from swelling and allows column packing. Protein adsorption was performed at different pH under static conditions. Furthermore, static capacities were determined. The material reveals high adsorption capacities for proteins under static conditions (270–408 mg/g), whereby proteins are mainly adsorbed via electrostatic interactions. The Freundlich isotherm is suggested as an adsorption model. For desorption a pH shift was found to be most effective. Binding and elution of human serum albumin and ovalbumin were tested under dynamic conditions. Dynamic capacities of about 40 mg/g for ovalbumin at 764 cm/h were found. The clay mineral provides suitable properties for the application as cost-efficient, alternative separation material.     

Adsorption studies in a synthetic rubber latex-ovalbumin system

1. Adsorption of ovalbumin on the latex surface was in excess of the quantity required to produce coverage of the surface over most of the protein concentrations range which was investigated. 2. "S" shaped isothermals which probably indicated multilayer adsorption were obtained. 3. The quantity of ovalbumin required to produce a constant surface charge density on the latex particle surface was a function of the pH, and a theory of active centers on the latex particles has been suggested. 4. A shift in the isoelectric point from that of native ovalbumin has been observed for the protein when adsorbed on a synthetic latex.     

Saliva as a medium for aldosterone measurement in repeated sampling studies

BackgroundSaliva is a readily available biological fluid, making it convenient in diagnosis of diseases and in multi-sampling protocols. Several salivary steroids give a useful index of free plasma levels. Increased incidence of primary aldosteronism (PA) in approximately 10% of the hypertensive population has increased interest in the mineralocorticoid aldosterone.MethodsA biotinylated-aldosterone tracer and a commercially available antibody are used in a time-resolved fluorescence immunoassay (TR-FIA) to measure salivary aldosterone (SA). Saliva was collected in various multi-sampling protocols: Investigation of diurnal rhythm in healthy and PA patients, ACTH stimulation test and posture test in healthy subjects.ResultsMethod validation showed a sensitivity of 19 ng/L and intra-/inter-assay precision between 7.2–10.1% and 8.7–15.7%, respectively. SA correlated significantly (y = 0.2995x ± 0.01, r² = 0.60) to plasma aldosterone measured by a commercial radioimmunoassay. SA (median; 95%CI) was at 111 (95–127) ng/L in PA (n = 84) and 50 (44–56) ng/L in healthy subjects (n = 60). After change in posture, aldosterone increased in both, saliva (57 (47–63) ng/L to 95 (84–117) ng/L) and plasma (26 (26–41) ng/L to 135 (110–181) ng/L). Peak levels were reached after 1 h, and were higher in females than in males.ConclusionsSA correlates well to plasma aldosterone and mirrors responses during conditions of stress. SA is significantly higher in PA, and the diurnal rhythm seen in the healthy is blunted in PA. We additionally found gender-dependent differential responses to posture, with higher increases in females. Measurement of aldosterone in saliva presents a useful and convenient method for application in multi-sampling studies.     

Adsorption of ligands on macromolecules in the fluctuating medium

In the present work fluctuations of number of ligands adsorbed on macromolecule are investigated. We have taken into account the adsorption and desorption of ligands under the circumstance of some adsorption centers fluctuations affected by medium fluctuation. The correlation function and spectral density of number of ligands adsorbed on macromolecule are calculated. The properties of these fluctuations which allow identifying a noisemaker are determined. It has been shown, thatfas andsluggis adsorption can be distinguished by properties of dispersion and spectral density. It has been also shown, that comparison of experimental and theoretical correlation functions (or spectral densities) allows to calculate constants of ligand - adsorption center binding and unbinding.     

Steroid bioconversion in a microemulsion system

The Delta(1,2)-dehydrogenation of high concentrations of the steroid -methyl-Reichstein's compound S-21-acetate (16MRSA) in a microemulsion system was studied using heat-dried and thawed Arthrobacter simplex cells as biocatalyst. The microemulsion system consists of an organic phase [75-95% (v/v)] with steroid (1-60 g/L(tot)), an aqueous phase [5-25% (v/v)] containing the cells (5-30 g/L(tot)), and a neutral surfactant (5-20 g/L organic solvent). Benzene derivatives, which solubilize 16MRSA up to 94 g/L, and phospholipids were used as organic solvents and surfactants, respectively, and menadione was added as an external electron acceptor. Factors affecting the dehydrogenation rate in the microemulsion system were studied. The influences of the 16MRSA and the menadione concentration on the dehydrogenation rate were described by Michaelis-Menten kinetics, apparent V'(max) and K'(m) values of 2.06 g/g dry weight h and 18.9 g/L for 16MRSA and 4.97 g/g dry weight h and 1.91 g/L for menadione being obtained. Optimal menadione concentration was dependent on the steroid concentration was dependent on the steroid concentration used. The reaction was strongly inhibited by high product concentrations. Much higher activities were obtained with the thawed cells than with the dried cells, conversions of 98% being reached within 14-16 h. for 16MRSA and cell dry weight concentrations of 40 and 10 g/L, respectively. Activity retention in a batch stirred tank reactor remained constant during the first 16-24 h of operation and then decreased, depending on the stirring rate; 22 to 65% of the initial reaction rate was obtained after 48 h at stirring rates of 650 and 2000 rpm, respectively.     

Pseudomonas: a promising biocatalyst for the bioconversion of terpenes

The Pseudomonas genus is one of the most diverse and ecologically significant groups of known bacteria, and it includes species that have been isolated worldwide in all types of environments. The bacteria from this genus are characterized by an elevated metabolic versatility, which is due to the presence of a complex enzymatic system. Investigations since the early 1960s have demonstrated their potential as biocatalysts for the production of industrially relevant and value-added flavor compounds from terpenes. Although terpenes are often removed from essential oils as undesirable components, its synthetic oxy-functionalized derivatives have broad applications in flavors/fragrances and pharmaceutical industries. Hence, biotransformation appears to be an effective tool for the structural modification of terpene hydrocarbons and terpenoids to synthesize novel and high-valued compounds. This review highlights the potential of Pseudomonas spp. as biocatalysts for the bioconversion of terpenes and summarizes the presently known bioflavors that are obtained from these processes.     

Field and laboratory studies on the bioconversion of coal to methane in the San Juan Basin

The bioconversion of coal to methane in the San Juan Basin, New Mexico, was investigated. Production waters were analyzed via enrichment studies, metabolite-profiling, and culture-independent methods. Analysis of 16S rRNA gene sequences indicated the presence of methanogens potentially capable of acetoclastic, hydrogenotrophic, and methylotrophic metabolisms, predominantly belonging to the Methanosarcinales and Methanomicrobiales. Incubations of produced water and coal readily produced methane, but there was no correlation between the thermal maturity and methanogenesis. Coal methanogenesis was greater when samples with a greater richness of Firmicutes were utilized. A greater archaeal diversity was observed in the presence of several aromatic and short-chain fatty acid metabolites. Incubations amended with lactate, hydrogen, formate, and short-chain alcohols produced methane above un-amended controls. Methanogenesis from acetate was not observed. Metabolite profiling showed the widespread occurrence of putative aromatic ring intermediates including benzoate, toluic acids, phthalic acids, and cresols. The detection of saturated and unsaturated alkylsuccinic acids indicated n-alkane and cyclic alkane/alkene metabolism. Microarray analysis complemented observations based on hybridization to functional genes related to the anaerobic metabolism of aromatic and aliphatic substrates. These data suggest that coal methanogenesis is unlikely to be limited by methanogen biomass, but rather the activation and degradation of coal constituents.     

Physiological studies of biosynthetic indole excretion in Bacillus alvei

Bacillus alvei excretes indole during early exponential growth in acid-hydrolyzed casein medium. l-Threonine is the amino acid responsible for "early" indole excretion, and the amount of indole excreted is directly related to the amount of l-threonine in the medium. "Early-indole" excretion can be prevented by the continuous addition of serine (3.1 mumoles per ml per hr) or by substituting a mutant with an impaired ability to degrade serine. The addition of serine to a culture during the period of indole excretion halts the excretion and stimulates indole utilization. Threonine is a competitive inhibitor of serine (K(i) = 0.6 m) in the tryptophan synthetase B reaction. The internal tryptophan concentration increases during the period of indole excretion, suggesting that threonine acts by increasing the activity of the tryptophan pathway. This view is supported by experiments demonstrating that anthranilic acid and indoleacrylic acid also stimulate indole excretion. A metabolic explanation is offered and discussed.     

Identification of indole compounds secreted byFrankia HFPArI3 in defined culture medium

Indole compounds secreted byFrankia sp. HFPArI3 in defined culture medium were identified with gas chromatography-mass spectrometry (GC-MS). WhenFrankia was grown in the presence of¹³C(ring-labelled)-L-tryptophan,¹³C-labelled indole-3-acetic acid (IAA), indole-3-ethanol (IEtOH), indole-3-lactic acid (ILA), and indole-3-methanol (IMeOH) were identified.High performance liquid chromatography (HPLC) and GC-MS with selected ion monitoring were used to quantify levels of IAA and IEtOH inFrankia culture medium. IEtOH was present in greater abundance than IAA in every experiment. When no exogenous trp was supplied, no or only low levels of indole compounds were detected.Seedling roots ofAlnus rubra incubated in axenic conditions in the presence of indole-3-ethanol formed more lateral roots than untreated plants, indicating that IEtOH is utilized by the host plant, with physiological effects that modify patterns of root primordium initiation.     

The separation of ferritin and haemosiderin for studies in the metabolism of iron

1. Ferritin and haemosiderin in diluted tissue cannot be quantitatively separated by centrifugation unless the pH is brought below 6. 2. These iron-storage substances can be separated from one another and from haemoglobin by ion-exchange chromatography on CM-cellulose, when good recoveries of total tissue iron are obtained. 3. The ferritin fraction has been identified by its solubility, its appearance under the electron microscope, crystallization with Cd²⁺ and the fact that it corresponds quantitatively to the heat-stable portion of the tissue non-haem iron.     

Engineering deacetoxycephalosporin C synthase as a catalyst for the bioconversion of penicillins

Journal of Industrial Microbiology & Biotechnology - 7-aminodeacetoxycephalosporanic acid (7-ADCA) is a key intermediate of many clinically useful semisynthetic cephalosporins that were...     

Thiophilic paramagnetic particles as a batch separation medium for the purification of antibodies from various source materials

A preparation of thiophilic agarose-based paramagnetic particles (T-Gel) has been developed with physical characteristics (particle size and particle density) that facilitate its use as a batch separation medium suitable for the large-scale purification and isolation of immunoglobulins. The medium was used to extract immunoglobulins from a wide range of starting materials, including sera, ascites fluid, tissue culture medium, and whole blood. None of these starting materials required pretreatment such as clarification by centrifugation or filtration prior to antibody extraction. The antibody purity obtained using T-Gel compared well with that obtained using protein A agarose column chromatography. Yields were approximately 30 mg of immunoglobulins per milliliter of T-Gel, and little was required in the way of specialist equipment. The method is uncomplicated and involves a roll mix extraction overnight, followed by magnetic separation to facilitate supernatant removal and subsequent washing of the particles. Elution of bound antibodies was carried out at neutral pH to yield a concentration of immunoglobulins that was approximately 7 mg/ml. The method was found to be applicable to antibody purification from the blood serum of seven different mammalian species and for all immunoglobulin classes.     

An unstructured model for studies on phenanthrene bioconversion by filamentous fungus Cunninghamella elegans

An unstructured model for phenanthrene bioconversion by filamentous fungus Cunnighamella elegans in stirred tank batch bioreactors was proposed. It was observed that the process of phenanthrene bioconversion is strictly associated with exponential growth of biomass. Therefore, a Monod-type, with Contois term, unstructured model could be applied to describe the process mathematically. The inhibition of biomass growth due to high concentrations of phenanthrene present in the medium was taken into account in the form of Yerusalimski's uncompetitive inhibition term.     

Steroid bioconversion in a novel aqueous two-phase system

A novel aqueous two-phase system, based on polyethyleneglycol (PEG) and monosodium glutamate, was tested for the 1-dehydrogenation of hydrocortisone-based substrates. This system led to higher substrate solubilities and biocatalyst/steroid separation levels when compared with alternative systems. The addition of short-chain monohydric alcohols resulted in higher solubilities and more favourable partition coefficients for the tested substrates. Bioconversion activities in PEG/glutamate systems with 2,5% (v/v) methanol were comparable to those measured in monophasic buffer-methanol medium.